The impact of anthelmintic treatment on human gut microbiota based on cross-sectional and pre- and postdeworming comparisons in Western Kenya

Murine studies suggest that the presence of some species of intestinal helminths is associated with changes in host microbiota composition and diversity. However, studies in humans have produced varied conclusions, and the impact appears to vary widely depending on the helminth species present. To demonstrate how molecular approaches to the human gut microbiome can provide insights into the complex interplay among disparate organisms, DNA was extracted from cryopreserved stools collected from residents of 5 rural Kenyan villages prior to and 3 weeks and 3 months following albendazole (ALB) therapy. Samples were analyzed by quantitative PCR (qPCR) for the presence of 8 species of intestinal parasites and by MiSeq 16S rRNA gene sequencing. Based on pretreatment results, the presence of neither Ascaris lumbricoides nor Necator americanus infection significantly altered the overall diversity of the microbiota in comparison with age-matched controls. Following ALB therapy and clearance of soil-transmitted helminths (STH), there were significant increases in the proportion of the microbiota made up by Clostridiales (P = 0.0002; average fold change, 0.57) and reductions in the proportion made up by Enterobacteriales (P = 0.0004; average fold change, -0.58). There was a significant posttreatment decrease in Chao1 richness, even among individuals who were uninfected pretreatment, suggesting that antimicrobial effects must be considered in any posttreatment setting. Nevertheless, the helminth-associated changes in Clostridiales and Enterobacteriales suggest that clearance of STH, and of N. americanus in particular, alters the gut microbiota.IMPORTANCE The gut microbiome is an important factor in human health. It is affected by what we eat, what medicines we take, and what infections we acquire. In turn, it affects the way we absorb nutrients and whether we have excessive intestinal inflammation. Intestinal worms may have an important impact on the composition of the gut microbiome. Without a complete understanding of the impact of mass deworming programs on the microbiome, it is impossible to accurately calculate the cost-effectiveness of such public health interventions and to guard against any possible deleterious side effects. Our research examines this question in a "real-world" setting, using a longitudinal cohort, in which individuals with and without worm infections are treated with deworming medication and followed up at both three weeks and three months posttreatment. We quantify the impact of roundworms and hookworms on gut microbial composition, suggesting that the impact is small, but that treatment of hookworm infection results in significant changes. This work points to the need for follow-up studies to further examine the impact of hookworm on the gut microbiota and determine the health consequences of the observed changes

Acquisition of pneumococci specific effector and regulatory Cd4+ T cells localising within human upper respiratory-tract mucosal lymphoid tissue

The upper respiratory tract mucosa is the location for commensal Streptococcus (S.) pneumoniae colonization and therefore represents a major site of contact between host and bacteria. The CD4(+) T cell response to pneumococcus is increasingly recognised as an important mediator of immunity that protects against invasive disease, with data suggesting a critical role for Th17 cells in mucosal clearance. By assessing CD4 T cell proliferative responses we demonstrate age-related sequestration of Th1 and Th17 CD4(+) T cells reactive to pneumococcal protein antigens within mucosal lymphoid tissue. CD25(hi) T cell depletion and utilisation of pneumococcal specific MHCII tetramers revealed the presence of antigen specific Tregs that utilised CTLA-4 and PDL-1 surface molecules to suppress these responses. The balance between mucosal effector and regulatory CD4(+) T cell immunity is likely to be critical to pneumococcal commensalism and the prevention of unwanted pathology associated with carriage. However, if dysregulated, such responses may render the host more susceptible to invasive pneumococcal infection and adversely affect the successful implementation of both polysaccharide-conjugate and novel protein-based pneumococcal vaccines

Signaling via interleukin-4, receptor alpha chain is required for successful vaccination against schistosomiasis in BALB/c mice

Radiation-attenuated (RA) schistosome larvae are potent stimulators of innate immune responses at the skin site of exposure (pinna) that are likely to be important factors in the development of Th1-mediated protective immunity. In addition to causing an influx of neutrophils, macrophages, and dendritic cells (DCs) into the dermis, RA larvae induced a cascade of chemokine and cytokine secretion following in vitro culture of pinna biopsy samples. While macrophage inflammatory protein 1 and interleukin-1 (IL-1) were produced transiently within the first few days, the Th1-promoting cytokines IL-12 and IL-18 were secreted at high levels until at least day 14. Assay of C3H/HeJ mice confirmed that IL-12 secretion was not due to lipopolysaccharide contaminants binding Toll-like receptor 4. Significantly, IL-12 p40 secretion was sustained in pinnae from vaccinated mice but not in those from nonprotected infected mice. In contrast, IL-10 was produced from both vaccinated and infected mice. This cytokine regulates IL-12-associated dermal inflammation, since in vaccinated IL-10/ mice, pinna thickness was greatly increased concurrent with elevated levels of IL-12 p40. A significant number of IL-12 p40 cells were detected as emigrants from in vitro-cultured pinnae, and most were within a population of rare large granular cells that were Ia, consistent with their being antigen-presenting cells. Labeling of IL-12 cells for CD11c, CD205, CD8, CD11b, and F4/80 indicated that the majority were myeloid DCs, although a proportion were CD11c F4/80, suggesting that macrophages were an additional source of IL-12 in the skin

Astragalus Polysaccharides Attenuate Postburn Sepsis via Inhibiting Negative Immunoregulation of CD4+CD25high T Cells

BACKGROUND: Astragalus polysaccharides (APS) isolated from one of the Chinese herbs, Astragalus mongholicus, are known to have a variety of immunomodulatory activities. However, it is not yet clear whether APS can exert an effect on the immune functions of regulatory T cells (Tregs). This study was carried out to investigate the effect of APS on the immune function of peripheral blood Tregs in postburn sepsis. METHODOLOGY/PRINCIPAL FINDINGS: BALB/C mice were randomly divided into six groups as follows: sham burn group, burn control (burn without infection animals) group, burn plus P. aeruginosa group, burn plus P. aeruginosa with APS (50 mg/kg) treatment group, burn plus P. aeruginosa with APS (100 mg/kg) treatment group, and burn plus P. aeruginosa with APS (200 mg/kg) treatment group, and they were sacrificed on postburn day 1, 3, 5, and 7, respectively, with seven animals at each time point. Magnetic microbeads were used to isolate peripheral blood Tregs and CD4(+) T cells. Phenotypes were analyzed by flow cytometry, and cytokine levels were determined with ELISA. In the burn plus P. aeruginosa group, forkhead/winged helix transcription factor p3 (Foxp3) expression on CD4(+)CD25(+)Tregs were strongly enhanced in comparison to the sham group, and the capacity of CD4(+)CD25(+)Tregs to produce interleukin (IL)-10 was markedly increased. Administration of APS to inhibit CD4(+)CD25(+)Tregs could significantly decrease expression of Foxp3 on CD4(+)CD25(+)Tregs, and IL-10 production in burned mice with P. aeruginosa infection. At the same time, proliferative activity and expression of IL-2 and IL-2Rα on CD4(+) T cells were restored. In contrast, anti-Toll-like receptor 4 (TLR4) antibody could block the effect of APS on Tregs immune function. CONCLUSION: APS might suppress CD4(+)CD25(+)Treg activity, at least in part, via binding TLR4 on Tregs and trigger a shift of Th2 to Th1 with activation of CD4(+) T cells in burned mice with P. aeruginosa infection

CD4+CD25+ T cells protect against experimentally induced asthma and alter pulmonary dendritic cell phenotype and function

The role of natural CD4+CD25+ regulatory T (T reg) cells in the control of allergic asthma remains poorly understood. We explore the impact of T reg cell depletion on the allergic response in mice susceptible (A/J) or comparatively resistant (C3H) to the development of allergen-induced airway hyperresponsiveness (AHR). In C3H mice, anti-CD25–mediated T reg cell depletion before house dust mite treatment increased several features of the allergic diathesis (AHR, eosinophilia, and IgE), which was concomitant with elevated T helper type 2 (Th2) cytokine production. In similarly T reg cell–depleted A/J mice, we observed a moderate increase in airway eosinophilia but no effects on AHR, IgE levels, or Th2 cytokine synthesis. As our experiments suggested that T reg cell depletion in C3H mice before sensitization was sufficient to enhance the allergic phenotype, we characterized dendritic cells (DCs) in T reg cell–depleted C3H mice. T reg cell–depleted mice had increased numbers of pulmonary myeloid DCs with elevated expression of major histocompatibility complex class II, CD80, and CD86. Moreover, DCs from T reg cell–depleted mice demonstrated an increased capacity to stimulate T cell proliferation and Th2 cytokine production, which was concomitant with reduced IL-12 expression. These data suggest that resistance to allergen-driven AHR is mediated in part by CD4+CD25+ T reg cell suppression of DC activation and that the absence of this regulatory pathway contributes to susceptibility

IL-10 from CD4+CD25−Foxp3−CD127− Adaptive Regulatory T Cells Modulates Parasite Clearance and Pathology during Malaria Infection

The outcome of malaria infection is determined, in part, by the balance of pro-inflammatory and regulatory immune responses. Failure to develop an effective pro-inflammatory response can lead to unrestricted parasite replication, whilst failure to regulate this response leads to the development of severe immunopathology. IL-10 and TGF-β are known to be important components of the regulatory response, but the cellular source of these cytokines is still unknown. Here we have examined the role of natural and adaptive regulatory T cells in the control of malaria infection and find that classical CD4+CD25hi (and Foxp3+) regulatory T cells do not significantly influence the outcome of infections with the lethal (17XL) strain of Plasmodium yoelii (PyL). In contrast, we find that adaptive IL-10-producing, CD4+ T cells (which are CD25−, Foxp3−, and CD127− and do not produce Th1, Th2, or Th17 associated cytokines) that are generated during both PyL and non-lethal P. yoelii 17X (PyNL) infections are able to down-regulate pro-inflammatory responses and impede parasite clearance. In summary, we have identified a population of induced Foxp3− regulatory (Tr1) T cells, characterised by production of IL-10 and down regulation of IL-7Rα, that modulates the inflammatory response to malaria

BALB/c Mice Infected with Antimony Treatment Refractory Isolate of Leishmania braziliensis Present Severe Lesions due to IL-4 Production

Leishmaniasis is a neglected disease that affects more than 12 million people worldwide. In Brazil, the cutaneous disease is more prevalent with about 28,000 new cases reported each year, and L. braziliensis is the main causative agent. The interesting data about the infection with this parasite is the wide variety of clinical manifestations that ranges from single ulcerated lesions to mucocutaneous and disseminated disease. However, experimental models to study the infection with this parasite are difficult to develop due to high resistance of most mouse strains to the infection, and the mechanisms underlying the distinct manifestations remain poorly understood. Here, the authors use a mouse experimental model of infection with different L. braziliensis isolates, known to induce diseases with distinct severity in the human hosts, to elucidate immune mechanisms that may be involved in the different manifestations. They showed that distinct parasite isolates may modulate host response, and increased IL-4 production and Arg I expression was related to more severe disease, resulting in longer length of disease with larger lesions and reduced parasite clearance. These findings may be useful in the identification of immunological targets to control L. braziliensis infection and potential clinical markers of disease progression

Immunity to Lutzomyia intermedia Saliva Modulates the Inflammatory Environment Induced by Leishmania braziliensis

Transmission of Leishmania parasites occurs during blood feeding, when infected female sand flies inject humans with parasites and saliva. Chemokines and cytokines are secreted proteins that regulate the initial immune responses and have the potential of attracting and activating cells. Herein, we studied the expression of such molecules and the cellular recruitment induced by salivary proteins of the Lutzomyia intermedia sand fly. Of note, Lutzomyia intermedia is the main vector of Leishmania braziliensis, a parasite species that causes cutaneous leishmaniasis, a disease associated with the development of destructive skin lesions that can be fatal if left untreated. We observed that L. intermedia salivary proteins induce a potent cellular recruitment and modify the expression profile of chemokines and cytokines in mice. More importantly, in mice previously immunized with L. intermedia saliva, the alteration in the initial inflammatory response was even more pronounced, in terms of the number of cells recruited and in terms of gene expression pattern. These findings indicate that an existing immunity to L. intermedia sand fly induces an important modulation in the initial immune response that may, in turn, promote parasite multiplication, leading to the development of cutaneous leishmaniasis

Induction of Protective CD4+ T Cell-Mediated Immunity by a Leishmania Peptide Delivered in Recombinant Influenza Viruses

The available evidence suggests that protective immunity to Leishmania is achieved by priming the CD4+ Th1 response. Therefore, we utilised a reverse genetics strategy to generate influenza A viruses to deliver an immunogenic Leishmania peptide. The single, immunodominant Leishmania-specific LACK158–173 CD4+ peptide was engineered into the neuraminidase stalk of H1N1 and H3N2 influenza A viruses. These recombinant viruses were used to vaccinate susceptible BALB/c mice to determine whether the resultant LACK158–173-specific CD4+ T cell responses protected against live L. major infection. We show that vaccination with influenza-LACK158–173 triggers LACK158–173-specific Th1-biased CD4+ T cell responses within an appropriate cytokine milieu (IFN-γ, IL-12), essential for the magnitude and quality of the Th1 response. A single intraperitoneal exposure (non-replicative route of immunisation) to recombinant influenza delivers immunogenic peptides, leading to a marked reduction (2–4 log) in parasite burden, albeit without reduction in lesion size. This correlated with increased numbers of IFN-γ-producing CD4+ T cells in vaccinated mice compared to controls. Importantly, the subsequent prime-boost approach with a serologically distinct strain of influenza (H1N1->H3N2) expressing LACK158–173 led to a marked reduction in both lesion size and parasite burdens in vaccination trials. This protection correlated with high levels of IFN-γ producing cells in the spleen, which were maintained for 6 weeks post-challenge indicating the longevity of this protective effector response. Thus, these experiments show that Leishmania-derived peptides delivered in the context of recombinant influenza viruses are immunogenic in vivo, and warrant investigation of similar vaccine strategies to generate parasite-specific immunity

Study of Leishmania pathogenesis in mice : experimental considerations

Although leishmaniases are endemic in 98 countries, they are still considered neglected tropical diseases. Leishmaniases are characterized by the emergence of new virulent and asymptomatic strains of Leishmania spp. and, as a consequence, by a very diverse clinical spectrum. To fight more efficiently these parasites, the mechanisms of host defense and of parasite virulence need to be thoroughly investigated. To this aim, animal models are widely used. However, the results obtained with these models are influenced by several experimental parameters, such as the mouse genetic background, parasite genotype, inoculation route/infection site, parasite dose and phlebotome saliva. In this review, we propose an update on their influence in the two main clinical forms of the disease: cutaneous and visceral leishmaniases